2' Modified oligonucleotides
a technology of oligonucleotides and modified oligonucleotides, which is applied in the field of modified oligonucleotides, can solve the problems of less than completely satisfactory, and achieve the effect of stable nucleas
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example 1
Preparation of Nucleotides and Oligomers (A=NHAc)
[0065]2′-N-acetaminouridine was protected at the 5′-position and made reactive at the 3′-position for formation of a phosphodiester linkage conjugating the residues in an oligomer as follows:
[0066]2′-N-Acylaminouridine. To 152 mg of 2′-N-acylamino-3′, 5′-O-diacyluridine (Verheyden, J. P. H, et al., Org Chem (1971) 36:250-254) (0.411 mmol) in 25 ml of MeOH was added a catalytic amount of KCN. After 15 h, 1.00 g of silica gel was added, and the reaction was concentrated. The powder was added to the top of a 20 mm column of silica gel that had been equilibrated in 5% H2O in CH3CN. The column was eluted with 5% H2O in CH3CN using standard flash chromatography conditions (Still, W. C., et al., J Org Chem (1978) 43:2923-2925). Isolation and concentration of the product afforded 59.5 mg (50.8% yield) of product.
[0067]2′-N-Acylamino-5′-O-(4,4′-dimethoxytrityl)-uridine. To 59.5 mg of 2′-N-acylaminouridine (0.208 mmol) in 2.5 ml of dry pyridine...
example 2
Preparation of Nucleotides and Oligomers
[0070]
[0071]2′-S-phenylcytidine was prepared from 2,2′-anhydro-(1-B-D-arabinofuranosyl) cytosine-HCl (a cyclic nucleoside) by suitable treatment with thiophenol. The —NH2 of cytosine and 5′-hydroxy were protected and the 3′—OH activated as follows:
[0072]2′-S-Phenylthiocytidine. To a solution of 500 mg (1.91 mmol) of 2,2′-anhydro-(1-B-D-arabinofuranosyl)-cytosine hydrochloride (purchased from Sigma) in 50 ml of dry DMF and 1.86 ml of dry TEA (13.3 mmol) was added 0.980 ml (9.54 mmol, 5.0 equiv) of thiphenol. The reaction was stirred for 5 h and then concentrated. The residue was concentrated from MeOH onto 2.00 g of silica gel. The powder was added to the top of a 30 mm column of silica gel that was equilibrated with CH2Cl2. The column was then eluted with one column volume of CH2Cl2, then one column volume of 6.25% MeOH in CH2Cl2, then one column volume of 12.5% MeOH in CH2Cl2 and then 25% MeOH in CH2Cl2. Concentration of the product fractions...
example 3
Preparation of Additional Nucleotides and Oligomers
A. (A=OCH2COOEt)
[0077]The 2′-derivatized nucleoside was prepared from N4-benzoyl-3′,5′-O-(tetraisopropyldisiloxane-1,3-diyl)-cytidine by reaction with ethyl iodoacetate and deprotection of the 2′ and 5′ hydroxyls. The 5′ position was protected and the 3′ position converted to a group reactive to link hydroxyl groups.
[0078]N4-Benzoyl-2′-O-(ethoxycaronylmethyl)-3′, 5′-O-(tetraisopropyldisiloxane-1,3-diyl)-cytidine. The preparation of this compound was an adaptation of a similar reaction used for the preparation of the 2′-OMe (Inoue, H., et al., Nucleic Acids Res (1987) 15:6131-6149). To 250 mg of N4-benzoyl-3′,5′-O-(tetraisopropyldisiloxane-1,3-diyl)-cytidine (Markiewicz, W. J., J Chem Res (1979) 181-196) (0.424 mmol; first concentrated from benzene) was added 1.00 ml (8.45 mmol, 19.9 equiv) of ethyl iodoacetate, followed by 294 mg of Ag2O (1.27 mmol, 2.99 equiv). The mixture was rapidly stirred and heated at 42° C. for 32 h. The mixt...
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